The repeated use of psychostimulants, opiates and alcohol produces long-term changes in brain chemistry and behavior that contribute to persistent and compulsive drug use. Our studies seek to understand the mechanisms, both at the systems and cellular level, which underlie these changes. Included in this goal is the identification of biological factors that may predispose certain individuals to initiate compulsive drug use and the identification of pharmacotherapies that may be effective in the treatment of drug and alcohol addiction. Our neurochemical studies suggest important and contrasting roles of endogenous opioid peptides (endorphins, enkephalins, dynorphins) and their receptors (mu: MOR, kappa: KOR; delta: DOR) in regulating neural transmission in those circuits within the brain upon which drugs of abuse act to affect and subsequently control behavior. Pharmacological inactivation or genetic ablation of MOR is associated with decreased basal release of dopamine in projection areas of mesolimbic dopamine neurons and a compensatory increase in the activity of the dopamine transporter; a membrane bound protein that regulates clearance of dopamine from the extracellular space. Similar alterations occur in response to genetic ablation of DOR. In contrast, KOR ablation leads to increased basal release of dopamine and a compensatory increase in the function of the dopamine transporter. Using behavioral and neurochemical approaches, we have shown that dysregulation of opioid peptide systems also leads to marked alterations in the responsiveness of an individual to several drugs of abuse. Pharmacological or genetic manipulations that selectively inactivate MOR abolish the effects of the prototypic opiate agonist, morphine. They also attenuate the behavioral effects of cocaine and alcohol. In contrast, KOR ablation results in increased sensitivity to the behavioral effects of cocaine, morphine and ethanol and the ability of these drugs to increase dopamine neurotransmission in the mesolimbic dopamine system is also enhanced. Using biochemical and immunocytochemical approaches we are examining the cellular mechanisms by which KOR ligands modulate dopamine neurotransmission and the responsiveness of dopamine neurons to cocaine. Studies in brain tissue have shown that acute KOR activation increases the activity of the dopamine transporter as well transporter number. Using biotinylation techniques we have obtained evidence that KOR activation is associated with increased trafficking of the dopamine transporter from the cytosol to the plasma membrane. Ongoing studies in HEK 293 cells that stably express fluorescently labeled KOR and dopamine transporters seek to determine whether increased trafficking also occurs in this model system and the cellular basis for this effect. We have also obtained evidence that KOR agonists may exert their effects via the activation of two kinase cascades (mitogen activated protein kinase and phosphoinositol -3 kinase) that regulate cytoskeletal dynamics. Increased activity of these cascades is associated with an increase in the activity and cell surface expression of the dopamine transporter in both synaptosomal preparation and heterologous expression systems whereas their inhibition decreases both transporter function and cell surface expression. We have also initiated studies examining the mechanisms by which dynorphin, the endogenous ligand for the KOR, can produce both opioid- and non-opioid receptor mediated effects. Our studies have shown that dynorphin can bind non-covalently to the NMDA receptor and facilitate NMDA-receptor evoked effects both in the brain and spinal cord. Furthermore, these non-opioid effects oppose those resulting from KOR activation. Studies conducted in-vivo and in-vitro indicate that the release of dynorphin and the ensuing facilitation of NMDA receptor activity contribute to the development of opiate tolerance as well to chronic pain resulting from nerve injury. In collaboration with IRP scientists, we have synthesized a decoy peptide that prevents the interaction of dynorphin with the NMDA receptor. The administration of this peptide attenuates the expression of morphine tolerance and the neurotoxic effects of dynorphin peptides. On-going studies are examining the utility of this peptide for the treatment of chronic pain, brain and spinal cord injury.